Pseudomonas aeruginosa (P. aeruginosa) is a leading cause of life-threatening nosocomial infections, especially in a compromised host. Human immunity to P. aeruginosa has been correlated with humoral antibody to lipopolysaccharide (LPS) and toxin A, as described in Pollack M. Huang A. I., Prescott R. K., Young L. S., Hunter K. W., Cruess D. F., Tsai C. M., "Enhanced survival in Pseudomonas aeruginosa septicemia associated with high levels of circulating antibody to Escherichia coli endotoxin core," J. Clin., Invest. 1983; 72: 1874-1881; Pollack M., Young, L. S., "Protective activity of antibodies to exotoxin A and lipopolysaccharide at the onset of Pseudomonas aeruginosa septicemia in man," J. Clin. Invest. 1979; 63: 276-286; and Cross A. C., Sadoff J. C., Iglewski B. H., Sokol P. A., "Evidence for the role of toxin A in the pathogenesis of infection with Pseudomonas aeruginosa in humans," J. Infect. Dis. 1980; 142: 538-546.
Anti-LPS antibody has been shown to be highly protective against P. aeruginosa infections in a variety of animal model systems, as noted in Cryz S. J. Jr., Furer E., Germanier R., "Protection against Pseudomonas aeruginosa infection in a murine burn wound sepsis model by passive transfer of antitoxin A, antilastase, and antilipopolysaccharide," Infect. Immun. 1983; 39: 1072-1079; Cryz S. J. Jr., Furer E., Germanier R., "Passive protection against Pseudomonas aeruginosa infection in an experimental leukopenic mouse model," Infect. Immun. 1983; 40: 659-664; Kazmierowski J. A., Reynolds H. Y., Kauffmann J. C., Durbin W. A., Graw R. G. Jr., Devlin H. B., "Experimental pneumonia due to Pseudomonas aeruginosa in leukopenic dogs: prolongation of survival by combined treatment with passive antibody to Pseudomonas and granulocyte transfusions," J. Infect. Dis. 1977; 135: 438-446; and Pier G. B., Sidberry H. F., Sadoff J. C., "Protective immunity induced in mice by immunization with high-molecular-weight polysaccharide from Pseudomonas aeruginosa," Infect. Immun. 1978; 22: 919-925. However, attempts to use native P. aeruginosa LPS as a vaccine have been hampered by a high frequency of adverse reactions following immunization and the need for numerous injections to evoke an optimal immune response, as noted in Alexander T. W., Fisher M., "Immunization against Pseudomonas infection after thermal injury," J. Infect. Dis. 1974; 130 (Suppl.): 152-158; Haghbin M., Armstrong D., Murphy M. L., "Controlled prospective trial of Pseudomonas aeruginosa vaccine in children with acute leukemia," Cancer 1973; 32: 761-766; and Young L. S., Meyer R. D., Armstrong D., "Pseudomonas aeruginosa vaccine in cancer patients," Annals Int. Med. 1973; 79: 518-527.
As described in Liu P.V., "Extracellular toxins of P. aeruginosa," J. Infect. Dis. 130 (Suppl.): 594-599 1974, toxin A is the most toxic product, on a weight basis, synthesized by P. aeruginosa. Toxin A acts to inhibit eucaryotic protein synthesis by catalyzing the transfer of the adenosine diphosphate-ribosyl (ADPR) moiety of nicotinamide adenine dinucleotide onto eucaryotic elongation factor 2, as discussed in Iglewski, B.H., Liu, P.V. and Kabat, D., "Mechanism of Action of P. aeruginosa exotoxin A: ADP-ribosylation of mammalian elongation factor 2 in vitro and in vivo," Infect. Immun. 15: 138-144, 1977 and Ohman, D.E., Burns R.P. and Iglewski B.H., "Corneal Infections in mice with toxin A and elastase mutants of P. aeruginosa," J. Infect. Dis. 142: 547-555, 1980. Antitoxin A antibody either passively administered or induced by active vaccination with a toxin A toxoid has provided significant protection against experimental P. aeruginosa infection. Additional investigations have demonstrated a direct correlation between antitoxic antibody and survival of patients from an episode of P. aeruginosa bacteremia, as described in Cross, A.S., Sadoff, J.C., Iglewski, B.H., and Sokol, P.A., "Evidence for the role of toxin A in the pathogenesis of human infection with Pseudomonas," J. Infect. Dis. 142: 538-46, 1980 and Pollack M.S. and Young, L.S., "Protective activity of antibodies to exotoxin A and lipopolysaccharides at the outset of P. aeruginosa septicemia in man," J. Clin. Invest. 63: 276-86, 1979.
Although native P. aeruginosa LPS contains protective serotype specific antigenic determinants, native LPS has been found to be too toxic for use in humans as a parenterally administered vaccine. Serotype specific antigenic determinants of P. aeruginosa are contained within the O-polysaccharide (PS) region of the LPS molecule, and although the PS can be isolated free of the toxic lipid A moiety of the LPS, the PS are non-immunogenic, as noted in Pier, G.B., Sidberry, H.F., and Sadoff, J.C., "Protective Immunity induced in mice by immunization with high molecular weight polysaccharide from P. aeruginosa," Infect. Immun. 22: 919-925, 1978 and Chester, I.R., Meadow, P.M., and Pitt, T.L., "The relationship between O-antigenic lipopolysaccharides and serological specificty in strains of P. aeruginosa of different O-serotypes," J. Gen. Microbiol, 78: 305-318, 1973. In order to induce a protective immune response to isolated PS, by the present invention isolated PS is covalently coupled to either tetanus toxoid or P. aeruginosa toxin A, which serve as "carrier proteins" for the PS. The conditions used to couple toxin A to PS effectively d-etoxify the toxin A molecule, thereby producing a PS-toxin A toxoid conjugate. The PS-toxin A and PS-tetanus toxoid conjugates are non-toxic, immunogenic and provide protection against experimental P. aeruginosa infections.